Our overall goal is to define the role(s) of a novel tolerogenic DC population in peripheral and central immune tolerance;to characterize the cellular and molecular mechanisms they employ to regulate T cell immunity;and to test their therapeutic potential in models of transplantation and immunity. Dendritic cells (DCs) present self or foreign antigen to T cells, and regulate immune responses. CCR9 expression defines a unique but widespread DC subset that migrates to the CCR9 ligand CCL25, a chemokine implicated in T progenitor cell homing to the thymus and in lymphocyte homing to the intestines. Preliminary studies show that CCR9+ DCs are a subset of immature plasmacytoid DC (pDC), and have provided initial insights leading to the following general hypotheses: 1. CCR9+ pDC act as potent endogenous tolerogens that induce peripheral regulatory T cells, and that can carry peripheral self antigens in a CCR9-dependent manner to the thymus where they may mediate central tolerance;and 2. CCR9+ pDC have the potential to suppress GVHD and allo-immune responses in transplantation. AIMS: 1: To further characterize CCR9+ pDC, confirm their tolerogenic potential, and define the cellular and molecular mechanisms CCR9+ pDCs employ to regulate T cell immunity. In vitro assays will confirm and assess mechanisms by which CCR9+ (vs. CCR9-) pDC induce Tregs, including regulatory cytokine expression and responses to TLR ligands. Comparative gene and protein expression analysis of CCR9+ (tolerogenic) vs CCR9- (immunogenic) pDC will lead to mechanistic insights into their specialized roles. In vivo approaches will confirm the ability of CCR9+ pDC to regulate immune responses in vivo. 2: To define the role of CCR9 in thymic homing and accumulation of tolerogenic pDCs. Short term homing and reconstitution assays will test our hypothesis that CCR9+ pDCs home better than CCR9- pDC to the thymus. The mechanistic importance of CCR9 to this homing will be defined with antibodies to the CCR9 ligand CCL25 and in comparative studies of pDC from wild type vs. CCR9-deficient donors. 3: To define the role of CCR9+ pDCs, and of pDC-expressed CCR9, in central tolerance. Established in vivo methods will define ability of thymic-homing CCR9+ pDC to regulate thymocytes development (clonal deletion, Treg induction), and the role of CCR9 in pDC mediated central tolerance. 4: To assess the therapeutic potential of CCR9+ DCs in models of tissue transplantation and to characterize their immunosuppressive mechanisms in vivo: Models of GVHD and pancreatic islet cell transplantation will be used to test the therapeutic potential of CCR9+ pDC mediated tolerogenesis, and to define the mechanisms involved. PUBLIC HEALTH RELEVANCE: These studies define an endogenous cell that appears to keep our immune system in check. They will test a novel approach using these cells for transplantation immunotherapy, and may lead to new insights and treatments for autoimmune, allergic and other disorders of the immune system.